MODIFIED POLYNUCLEOTIDES FOR THE PRODUCTION OF PROTEINS
The invention relates to compositions and methods for the preparation, manufacture and therapeutic use of polynucleotides, primary transcripts and mmRNA molecules.
1 . An isolated polynucleotide comprising;
(a) a first region of linked nucleosides, said first region encoding a polypeptide of interest, said polypeptide of interest selected from the group consisting of SEQ ID NOs 8922-17687;
(b) a first flanking region located at the 5′ terminus of said first region comprising;
(i) a sequence of linked nucleosides selected from the group consisting of the native 5′ UTR of any of the nucleic acids that encode any of SEQ ID NOs 8922-17687, SEQ ID NOs: 1-4 and functional variants thereof; and
(ii) at least one 5′ terminal cap;
(c) a second flanking region located at the 3′ terminus of said first region comprising;
(i′) a sequence of linked nucleosides selected from the group consisting of the native 3′ UTR of any of the nucleic acids that encode any of SEQ ID NOs 8922-17687, SEQ ID NOs 5-21 and functional variants thereof; and
(ii′) a 3′ tailing sequence of linked nucleosides.
2 . The isolated polynucleotide of claim 1 wherein the first region of linked nucleosides comprises at least an open reading frame of a nucleic acid sequence, wherein the nucleic acid sequence selected from the group consisting of SEQ ID NOs: 17688-61633.
3 . The isolated polynucleotide of claim 1 , wherein the 3′ tailing sequence of linked nucleosides is selected from the group consisting of a poly-A tail of approximately 160 nucleotides and a polyA-G quartet.
4 . The isolated polynucleotide of claim 1 which is purified.
5 . The isolated polynucleotide of claim 1 , wherein the at least one 5′ terminal cap is selected from the group consisting of Cap0, Cap1, ARCA, inosine, N1-methyl-guanosine, 2′fluoro-guanosine, 7-deaza-guanosine, 8-oxo-guanosine, 2-amino-guanosine, LNA-guanosine, and 2-azido-guanosine.
6 . The isolated polynucleotide of claim 1 , wherein at least one of said linked nucleosides comprises at least one modification as compared to the chemical structure of an A, G, U or C ribonucleotide.
7 . The isolated polynucleotide of claim 6 , wherein at least one said modification is located in a nucleoside base and/or sugar portion.
8 - 37 . (canceled)
38 . A pharmaceutical composition comprising the isolated polynucleotide of claim 1 and a pharmaceutically acceptable excipient.
39 - 40 . (canceled)
41 . A method of producing a polypeptide of interest in a mammalian cell, tissue or organism comprising administering to said cell, tissue or organism the pharmaceutical composition of claim 38 .
42 . The method of claim 41 , wherein the isolated polynucleotide is formulated.
43 . The method of claim 42 , wherein the formulation comprises a lipid.
44 - 50 . (canceled)
51 . A method for producing an increased level of a polypeptide of interest selected from the group consisting of SEQ ID NOs 8922-17687 in a mammalian cell, tissue or organism, comprising administering to said cell, tissue or organism a total daily dose of the pharmaceutical composition of claim 38 in two or more equal or unequal split doses.
52 - 76 . (canceled)